The redox-sensitive R-loop of the carbon control protein SbtB contributes to the regulation of the cyanobacterial CCM

Abstract

SbtB is a PII-like protein that regulates the carbon-concentrating mechanism (CCM) in cyanobacteria. SbtB proteins can bind many adenyl nucleotides and possess a characteristic C-terminal redox sensitive loop (R-loop) that forms a disulfide bridge in response to the diurnal state of the cell. SbtBs also possess an ATPase/ADPase activity that is modulated by the redox-state of the R-loop. To investigate the R-loop in the cyanobacterium Synechocystis sp. PCC 6803, site-specific mutants, unable to form the hairpin and permanently in the reduced state, and a R-loop truncation mutant, were characterized under different inorganic carbon (Ci) and light regimes. Growth under diurnal rhythm showed a role of the R-loop as sensor for acclimation to changing light conditions. The redox-state of the R-loop was found to impact the binding of the adenyl-nucleotides to SbtB, its membrane association and thereby the CCM regulation, while these phenotypes disappeared after truncation of the R-loop. Collectively, our data imply that the redox-sensitive R-loop provides an additional regulatory layer to SbtB, linking the CO2-related signaling activity of SbtB with the redox state of cells, mainly reporting the actual light conditions. This regulation not only coordinates CCM activity in the diurnal rhythm but also affects the primary carbon metabolism.

The second messenger c-di-AMP controls natural competence via ComFB signaling protein

“…Binding of recombinantly produced ComFB to c-di-AMP was analysed in vitro by isothermal titration calorimetry (ITC), thermal shift assay (TSA), and differential scanning fluorimetry (nanoDSF), as described previously (Lapina et al 2018; Haffner et al 2023a; Mantovani et al 2023b). For ITC and TSA, both ComFB and c-di-AMP were dissolved in binding buffer (50 mM Tris/HCl, pH 8.0, 300 mM NaCl, 0.55 mM EDTA).…”

Cyanobacteria form a procarboxysome-like structure in response to high CO2

“…11,20,24 However, carboxysomal redox state has neither been directly compared to the cytosol nor has a specific redox pool been targeted, 11,20,24 so much remains to be explored on the redox relationship of the carboxysome to the cytosol under variable conditions. The activity and function of several CCM proteins are known to be redox-regulated, such as one of the HCO3membrane transporters, SbtB/A, as way to modulate carbon uptake, 19,25 and the scaffold protein, CcmM, as a way to adjust RuBisCO packing during carboxysome formation. 16,17,[20][21][22] Others have been indicated as redox sensitive, such the shell protein, CcmK4, 21,22 and both the large and small subunits of RuBisCO, but it is unknown what these redox sensitivities achieve.…”

Diurnal rhythm causes metabolic crises in the cyanobacterial mutants of c-di-AMP signalling cascade

“…Therefore, night phases require a metabolic switch to heterotrophic-like carbon catabolism, which includes glycogen catabolism mainly via the oxidative pentose-phosphate (OPP) pathway (Gründel et SbtB, a small regulatory protein belonging to the PII signaling superfamily (Forchhammer et al 2022), is encoded in an operon together with the low Ci-induced sodium-dependent bicarbonate transporter SbtA (Selim et al 2018, Forchhammer andSelim 2020). Similar to canonical PII proteins (Lapina et al 2018, Selim et al 2019& 2020a, SbtB binds the adenine nucleotides ATP and ADP, but in contrast to these, it also binds AMP, cAMP and c-di-AMP (Selim et al 2018(Selim et al & 2021, thus combining the cell's energy status with second messenger signaling, which directly influences the SbtA-B complex formation (Selim et al 2023) and many other processes related to Ci acclimation (Mantovani et al 2022(Mantovani et al , 2023a(Mantovani et al & 2023b. Under low Ci conditions, intracellular AMP levels rise, thus leading to a strong SbtA-B complex in which SbtB serves as plug-like structure to prevent HCO3 -leakage (Haffner et al 2023.…”